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16, 235-239 (1979) by Springer-Verlag 1979 Effect of Oral Galactose on GIP and Insulin Secretion in Man M. Morgan, J. W. Wright, and V. Marks Department of Biochemistry, Division of Clinical Biochemistry, University of Surrey, Guildford, England insulinotropic effect of 50 g galactose given orally to 5 normal volunteers on two occasions - once with and once without a period of hypergly- caemia produced by an intravenous glucose infusion - was studied. Oral galactose caused a words: oral galactose, insulin secretion, enteroinsular axis, induced hyperglycaemia. Glucose stimulates insulin secretion by direct action on the B-cell as well as by the liberation of intestinal insulin releasing hormones. Galactose, on the other hand, though capable of stimulating insulin secretion when taken orally 8 fails to do so when adminis- tered intravenously 6, 8. This suggests that the insu- and Methods healthy volunteers drawn from amongst the laboratory staff participated in the experiments. Three were men and two were women and their ages ranged between 25 and 35 years. All were within 10% of ideal body weight and had been eating a normal diet containing not less than 200 g carbohydrate per day until the even- ing preceding the tests. None of the volunteers were taking any drugs at the time of study. Each subject attended the laboratory on two occasions having fasted since 2000 h 0012-186X/79/0016/0235/$01.00 L. M. Morgan et al.: Effect of Oral Galactose on GIP and Insulin Secretion 1,000 Glucose (0.25g/kg/hr IV) 8OO v Glucose Oral Galaetos "', (50g) 200 ! 60 90 120 ! 180 Effects of the ingestion of galactose either with or without intravenous glucose infusion on plasma immunoreactive GIP, in 5 normal subjects. (Q--O = galactose alone 0 ..... 0 = galactose + i. v. glucose, ~ = mean _ S. E. M. *p ) 50 40 30 z- 20 (0.25g/kg/hr IV) ' Glucose T, I 5g) (O;gl) Galactos,e~' 1, ''I~'',T 0 90 ,0 150 ,80 huin) 2. Effects of the ingestion of galactose either with or without intravenous glucose infusion on plasma immunoreactive insulin, in 5 normal subjects. (0--0 = galactose alone, o ..... o = galactose + i. v. glucose, = mean + S. E. M.) ple was put into lithium heparin tubes containing sufficient aproti- nin to produce a final concentration of 500 k. i. units/ml blood. The mixture was centrifuged immediately, the plasma separated, frozen and stored at -20 ~ until assayed for immunoreactive insulin 14 and GIP 10. Measurement of GIP was by radioimmunoassay using a rabbit antiserum raised against porcine GIP, using porcine G1P stand- ards. The assay was able to distinguish 70-110 pg/ml GIP from zero in plasma samples. The antiserum showed less than 1% cross reactivity with glucagon, GIP, secretin, pancreatic polypeptide, insulin or C-peptide. Preliminary work indicates that the anti- serum can recognise big GIP (MW 8,000 Daltons) and standard GIP (MW 5,300 Daitons). (O'Do

risio, T., Personal communica- tion). The "galactose-only" tests were carried out first; the infusion experiments later. The subjects sat comfortably, at ease, in the investigation unit throughout their performance. None of the sub- jects experienced abdominal or other discomfort at any time dur- ing the tests. The results were compared using Student's paired-t test. Informed consent was obtained from all subjects, and permis- sion to carry out the experiments was obtained from St. Luke's Hospital ethical committee. M. Morgan et al.: Effect of Oral Galactose on GIP and Insulin Secretion Glucose (0.25g/kg/hr IV) 2.0 S 1.5 v ~1.0 0.5 *#  )ral Galactose 2 I I i I I I I I 90 120 150 180 Time (mm) Fig. 3. Plasma galactose levels attained following the ingestion of 50 g galactose either with or with- out intravenous glucose infusion, in 5 normal sub- jects. (0--0 = galactose alone, o ..... o = galac- tose + i. v. glucose, ~ = mean _+ S. E. M., *p **p ) ~8 2 6 IV Glucose (12.5g) Glucose (0.25g/kg/hr IV) Galactose '~. (50g) i ! I I I l 30 60 90 120 150 180 Time (min) 4. Effects of the ingestion of galactose either with or without intravenous glucose infusion on blood glucose, in 5 normal sub- jects. (0--0 = galactose alone, o ..... o = galactose + i. v. glucose, ~ = mean + S. E.M.) results of the two series of experiments are sum- marised and compared in Figures 1-4. Galactose Alone ingestion of galactose was followed in every case (Fig. 1) by a large and rapid rise in plasma GIP (mean peak level 900 _+ 65 ng/1 at 30 min) and a small (Fig. 2) rise in plasma insulin levels. Plasma galactose levels rose slowly at first, reaching a peak (mean peak level 1.97 _+ 0.28 mmol/1) at 60 min and had returned almost to basal values by 120min (Fig. 3). Blood glucose showed a small but significant rise during the first 45 min following the ingestion of galactose (Fig. 4) but had returned to baseline values, or below, by 90 min. L.M. Morgan et al.: Effect of Oral Galactose on GIP and Insulin Secretion Glucose Plus Oral Galactose injection, followed by continuous infusion of glucose, produced a rapid rise both in blood glucose and plasma insulin levels (Figs. 2 & 4). Blood glucose levels reached a peak within five minutes of the bolus injection and fell slowly thereafter despite constant intravenous infusion of glucose at the rate of 0.25 g/ kg/h (Fig. 4). Plasma insulin levels rose rapidly ini- tially (Fig. 2) but fell almost equally rapidly to a new supra-basal plateau. Thereafter, plasma insulin levels remained more or less constant until the ingestion of oral galactose when they rose once again, this time to a level considerably higher than after galactose alone (53.3 _+ 10.7 mU/1 v. 20.5 + 2.8 mU/1). Comparison of the total area under the plasma insulin curve fol- lowing the administration of oral galactose (from 60-180min) showed a significantly greater insu- li

naemic response to galactose given during the IV glucose infusion than when galactose was given alone. (Area under curve with galactose alone, 236.5 + 66.0, with galactose + IV glucose 451.9 _+ 81.6, p 0.025 estimated using the 60 min plasma insulin level as a baseline). Plasma GIP levels declined slowly and signifi- cantly throughout the period of intravenous glucose infusion until interrupted by the ingestion of galac- tose (GIP levels between 10 and 60 min being sig- nificantly lower (p 0.05) than plasma GIP at zero time). Thereafter, plasma GIP levels rose (mean peak level 595 _+ 110 ng/1 at 45 minutes) though more slowly and to a significantly lesser extent than after oral galactose alone. Plasma galactose levels (Fig. 3) showed a consistently smaller rise (p 0.025) than when galactose was ingested in the ab- sence of an intravenous glucose infusion. Discussion The work reported here indicates that, in man, galac- tose ingestion, like that of glucose 3 is associated with a large brisk rise in plasma GIP concentrations. Reports of the GIP releasing properties of galactose are conflicting. Of the three abstracts relating to GIP release in dogs after oral galactose, two report a rise in plasma GIP levels - albeit a rather small one - and one no significant change 9, 16, 18. One recent communication 13 relating to man reports a small rise in plasma GIP levels after oral galactose. GIP is relatively ineffective in stimulating insulin release except in the presence of mild to moderate hyperglycaemia 3. This may account for some of the reported failures to observe a rise in plasma insulin levels following oral galactose administration 1. Since the insulinotropic effect of GIP is enhanced by hyperglycaemia we decided to compare the GIP releasing and insulinotropic effects of oral galactose in the presence and absence of induced hypergly- caemia. It did not prove possible, as had been intended, to maintain a constant blood glucose con- centration throughout the period of induced hyperglycaemia. Nevertheless, even in the presence of a declining blood glucose concentration the administration of 50g galactose by mouth was promptly followed by a large rise in plasma insulin concentration. A small rise in blood glucose concentration is commonly, though not invariably, observed during the first hour following ingestion of galactose as in the present series of experiments. We suggest that this small rise in blood glucose concentration makes the B-cells of the pancreas receptive to the rise in plasma GIP levels produced by oral galactose and indirectly responsible, therefore, for the modest rise in plasma insulin concentration observed when galac- tose is ingested alone 8, 15. The much larger rise in plasma insulin provoked by the ingestion of galactose in the artificially induced hyperglycaemic subjects was associated with a significantly smaller and delayed rise in plasma

GIP concentration. The reduc- tion in plasma GIP response to oral galactose under these circumstances could be accounted for by opera- tion of a negative feedback control of GIP secretion by insulin 3. It could, however, be due to inhibition of galactose absorption. The much lower plasma galactose level observed when the subjects drank their galactose solution whilst receiving IV glucose compared with when they drank it alone, was unexpected. Addition of glucose into the gut has been reported to interfere with active jejunal galactose transport to different degrees in various species 5, 12. It appears unlikely that IV glucose could act in this way, although the possibility must be considered that hyperglycaemia in some way delays gastric emptying - possibly by inhibition of motilin secretion (Bloom, S. R., personal communi- cation). Beyreiss and co-workers 2 reported the inhibition of galactose absorption from the gut in rabbits by the infusion of exogenous insulin. The higher endogenous insulin levels attained when our human subjects received IV glucose could therefore interfere with their galactose absorption. Another explanation of the lower rise in plasma galactose levels when subjects were receiving IV glucose is that whilst galactose absorption is unchanged by this ma- noeuvre, its clearance from the plasma is increased as a result of the greater hyperinsulinaemia. Most cells of the body - with the notable exception of those in the liver - are virtually impermeable to glucose and M. Morgan et al.: Effect of Oral Galactose on GIP and Insulin Secretion 239 except in the presence of insulin 4, 7, 17. Oral galactose alone is a poor stimulus to insulin se- cretion and consequently galactose is excluded from entry into all but liver cells. However, the higher insulin levels attained when subjects received IV glu- cose could render peripheral tissues more permeable to galactose. Animal experiments have shown that exogenous insulin increases galactose removed from the blood and its entry into peripheral tissues 6, 7, 17 where it is probably metabolised by a pathway not involving the enzyme galactokinase. The role of insulin is regulating the metabolism of galactose has largely been ignored in the past, prob- ably because, under most experimental conditions it does not stimulate insulin secretion - or does so to only a slight extent. Under most natural conditions of usage, however, namely when consumed as lactose, galactose can and does stimulate insulin secretion which could then proceed to play a part in its metabolism. Ambrus, J. L., Ambrus, C. M., Shields, R., Mink, I. B., Cleve- land, C.: Effect of galactose and sugar substitutes on blood insulin levels in normal and obese subjects. J. Med. 7, 429-432 (1976) 2. Beyreiss, K., Muller, F., Strack, E.: Iber die Resorption von Monosacchariden. I. Der EinfluB von Insulin auf die Resorp- tion der Galaktose. Z. Ges. Ex

p. Med., 138, 277 (1964) 3. Brown, J. C., Dryburgh, J. R., Ross, S. A., Dupr6, J.: Identifi- cation and actions of gastric inhibitory polypeptide. Recent Prog. Horm. Res. 31, 487-532 (1975) 4. Goldstein, M. S., Henry, W.L., Huddlestun, B., Levine, R.: Action of insulin on transfer of sugars across cell barriers: common chemical configuration of substances responsive to action of the hormone. Am. J. Physiol. 173, 207 (1953) 5. Holdsworth, C. D., Dawson, A. M.: The absorption of mono- saccharides in man. Clin. Sci. Mol. Med. 27, 371-379 (1964) 6. Kvam, D. C.: Lack of effect of tolbutamine upon certain insu- lin-responsive sugars. Proc. Soc. Exp. Biol. Med. 115, 904-906 (1964) 7. Levine, R., Goldstein, M.S., Huddlestun, B., Klein, S.P.: Action of insulin on the "permeability" of cells to free hexoses as studied by its effect in the distribution of galactose. Am. J. Physiol. 163, 70-83 (1950) 8. Marks, V., Samols, E.: Action de diff6rents stimuli (galactose, fructose, et lipides) sur l'insulin s6crdtion humaine: influences du tractus gastro-intestinal. J. Annu. Diabetol. H6tel Dieu 9, 179-190 (1969) 9. Martin, E.W., Sirinek, K.R., Crockett, S.E., O'Dorisio, T.M., Mazzaferri, E.L., Thomford, N.R., Cataland, S.: Release of gastric inhibitory polypeptide: comparison of hyperosmolar carbohydrate solutions as stimuli. Surg. Forum 26, 381-382 (1975) 10. Morgan, L. M., Morris, B. A., Marks, V.: Radioimmunoassay of gastric inhibitory polypeptide. Ann. Clin. Biochem. 15, 172-177 Morley, G., Dawson, A., Marks, V.: Manual and autoanalyser methods for measuring blood glucose using guaicum and glu- cose oxidase. Ann. Clin. Biochem. 5, 42-45 (1968) 12. Ricklis, E., Haber, B., Quastel, J. H.: Absorption of mixtures of sugars by isolated surviving guinea pig intestine. Can. J. Biochem. 36, 373-380 (1958) 13. Ross, S.A., Dupr6, J.: Effects of ingestion of triglyceride or galactose on secretion of gastric inhibitory polypeptide and on responses to intravenous glucose in normal and diabetic sub- jects. Diabetes 27, 327-333 (1978) 14. Samols, E., Bilkus, D. A.: A comparison of insulin immunoas- says. Proc. Soc. Exp. Biol. Med. 115, 79-84 (1964) 15. Shima, K., Kuroda, K., Matsuyama, T., Tarui, S., Nishikawa, M.: Plasma glucagon and insulin responses to various sugars in gastrectomized and normal subjects. Proc. Soc. Exp. Biol. Med. 139, 1042-1048 (1972) 16. Sirinek, K. R., Thomford, N. R., Pace, W. G., Crockett, S. E., O'Dorisio, T. M., Mazzaferri, E. L., Cataland, S.: J. Surg. Res. (in press) (1978) 17. Wick, A.N., Drury, D.R.: Action of insulin on volume of distribution of galactose in the body. Am. J. Physiol. 173, 229-232 (1953) 18. Williams, R.H., Biesbroek, J.: Effect of single food compo- nents on plasma GIP. Diabetes 26, 374 (1977) Received: July 28, 1978 and in revised form: November 28, 1978 Dr. L. M. Morgan Department of Biochemistry University of Surrey Guildford, GU2 5XH Eng